The objective of this application is to (I) design and synthesize photoresponsive [60]fullerenyl enantiomers and related nanostructures as photosensitizers for PDT applications and (ii) carry out in vitro and in vivo biostudies of amphiphilic multifunctional [60]fullerenyl enantiomers and chromophore nanostructures as 1PA-PDT and 2PA-PDT (2-3 absorptive) nanomedicines. We propose to design and synthesize novel cationic stereoregular 1-pyrrolino[60]fullerenyl enantiomers PF-PhOEGn and cationic C60(>CPAF-EG6)x for the use as PDT agents with the following characteristics: (1) high photo stability for multiple PDT treatments with only a single-dose of the drug; (2) enhancement of light-harvesting capability using the C60-antenna construction, for cytotoxic ROS production; (3) capability of molecular self-assembly of C60-antenna forming stable vesicles for high volume delivery of PDT drug; (4) large enhancement of two-photon absorption cross-sections [>4500 GM for C60- (antenna)2] for effective 2PA-PDT treatments. We have established some of the molecular structural features of fullerenes that mediate broad-spectrum photokilling of microbial cells and cancer cells. This insight into required structural motifs has been applied to the molecular design of fullerene nanomedicines. We will study PDT efficiency by both in vitro and in vivo experiments: (1) we have assembled a panel of pathogenic microorganisms designed to encompass most of the microbes that cause localized infections e.g. in wounds and burns, and which demonstrate both intrinsic and acquired multi-antibiotic resistance. Polycationic fullerenes will be tested as PDT agents against these pathogens. (2) For fullerene-mediated PDT treatment against localized infections we will study a mouse model of wound infection using genetically engineered bioluminescent bacteria combined with optical imaging to follow the progress of the infection non-invasively in real time. (3) We will carry out in vitro PDT against a panel of mouse cancer cell lines that are expected to have different uptakes of fullerenes and different cellular responses to reactive oxygen species produced by PDT. Dose response studies of both drug and light will be carried out for both 1-photon and 2-photon excitation and mechanisms of cell death studied. (4) For anti-cancer testing we will use subcutaneous mouse tumors and compare intratumoral injection with IV delivery of the fullerene. We will compare both 1-photon excitation using a KTP laser and 2-photon excitation using a femtosecond Ti:sapph laser. PUBLIC HEALTH RELEVANCE: The fullerene nanomedicines described in this application have the potential to mediate photodynamic therapy of two of the major killer diseases of the present age. Multi-drug resistant bacterial infections and cancerous tumors could be eliminated by a combination of the appropriate molecular design of fullerene and illumination with CW or femtosecond pulsed light. Particular chemical and photophysical features of these compounds proposed herein will improve PDT of these diseases beyond what is presently possible.